Can we understand what triggered
the separation of the two ancestral lineages that led to human and chimpanzees
today? Although it happened so long ago, scientists are homing in on the
process – one which contains powerful explanatory evidence that brings a further
level of detailed understanding to Human Evolution.

The most famous speciation
of all

Around six million years
ago, the most famous speciation of all occurred – the divergence of the lineage
that eventually led to modern humans from the lineage that led to chimpanzees
(1) (2) (3). That speciation initiated the evolutionary process that led
to the divergence of mankind from our nearest living cousin, the chimpanzee.

There is a long-standing
controversy amongst palaeontologists who debate whether modern humans evolved
in Africa (the Out of Africa hypothesis), or arose simultaneously in many different
places in the world after human ancestors had migrated out of Africa (the multi-regional
hypothesis) (5). Recent molecular data tend to favour the Out of Africa
model (genomic diversity is greater amongst Africans than populations of other
continents and ranges – depending on the specific parts of the genome considered
and the actual methods employed – from marginally to greatly more diverse).
Although recent data support the Out of Africa hypothesis, it is not a
simple, straightforward picture and there seems to have been considerable migrations
into and out of Africa and Asia of Homo erectus.

However, these controversies
focus on the later stages of the evolution of Homo sapiens. There is little
or no disagreement that the common ancestor of humans and chimpanzees lived in Africa
and that the original divergence of the two lineages took place in that continent.
So whatever the more recent human evolutionary history might be, we can ask
detailed questions about what caused the divergence of the two lineages from
their common ancestor.

What led to the divergence
of man and chimpanzee lineages?

So, if the common ancestor
of man and chimpanzee lived in Africa six million years ago, what led to the
genetic isolation between parts of the population that resulted in two species
developing – species that were first cousins and that were the ancestors of
the two lineages that eventually evolved to form modern humans and modern chimpanzees?

One hypothesis for this
mechanism is that part of the population of the common ancestor became geographically
isolated from the rest of the population over a long period of time, perhaps
owing to a barrier such as the Rift Valley, or mountains or rivers. Such
geographic isolation would prevent the process of gene flow from the population
on one side of the barrier to the population on the other side. Over time,
different mutations would accumulate in the two populations, resulting in different
evolutionary paths and ultimately in an inability to interbreed. Once populations
no longer interbreed in the wild, there can be no gene flow between them and
each will follow its own divergent evolutionary path. This process is known
as allopatry.

Another hypothesis is that
different subgroups of the same species, for some reason such as sexual preference
or specialisation to a particular narrow niche, stop interbreeding and eventually
become incapable of doing so – speciation results but without geographic separation.
This is known as sympatric speciation.

One type of sympatric speciation
is potentially caused by major mutations that prevent successful interbreeding
between parts of the population that have the mutation and parts that do not:
mating between the two groups either does not result in offspring or in hybrid
offspring which are themselves sterile (whilst breeding within each group –
that with and that without the major mutation – is fully fertile).

It would be interesting
to know which of these mechanisms led to the divergence of human and chimpanzee
lineages all those millions of years ago.

Chromosomal rearrangements

So what kind of major mutation
could set up a barrier to fertile mating and gene flow between subgroups within
what were originally the same species? One possibility is chromosomal
rearrangements. Chromosomal rearrangements occur when substantial tracts
of DNA are inverted or repositioned on the chromosome. Chromosomal rearrangements
that fix in the genome are relatively common. Comparison of the mouse
and human genomes indicate that about 300 rearrangements have occurred since
the divergence of mouse and human lineages – that is about one rearrangement
every 200,000 years (5).

In the case of human and
chimpanzee, the divergence occurred sufficiently long ago that there are significant
numbers of rearrangements in the chromosomes but not so long that all chromosomes
have rearrangements. Chromosomes 1, 4, 5, 9,12, 15, 16, 17 and 18 in humans
have inversions of major tracts of code compared with homologous chromosomes
in chimpanzees, and human chromosome 2 results from the end to end fusion of
two acrocentric chromosomes that remain separate in all the other great apes.
(6), (7), (8). Go here for a more comprehensive explanation of chromosome
2 fusion.

Could it be that some or
all of these chromosomal rearrangements were responsible for triggering the
speciation that separated the two lineages?

There is a simple model
for the idea that chromosomal rearrangements can lead to reproductive isolation
(even in geographically contiguous populations). The classical concept
is that individuals that are heterozygous for the rearrangement (ie individuals
who carry one copy of the chromosome that has the rearrangement mutation and
one that has not) are less fertile than homozygous individuals (ie those who
carry both copies of the chromosome either rearranged or in the original unmodified
form). This is caused because recombination between the two copies of the
chromosome in this case would result in duplication and deletion of substantial
amounts of genetic code. Selection would then favour the homozygous
individuals and this would lead to separation of the population into two separate
homozygous subpopulations, one with and one without the chromosomal rearrangement.
Those subpopulations would be, in effect, reproductively isolated, and
this would eventually lead to speciation. So, as this model goes, the
random mutational event of a chromosomal rearrangement can be the trigger and
cause for speciation.

However, there is a major
paradox with this hypothesis that calls its validity into question. If
heterozygous individuals are much less fertile than normal homozygous individuals,
it is hard or impossible to see how the rearrangement mutation gets established
in a sub-population at all. It would simply be selected out. If,
on the other hand, heterozygous individuals are as fertile or almost as fertile
as homozygous individuals, the rearrangement will have no more difficulty in
becoming established than any other neutral mutation. And it would present
no reproductive barrier. Populations would exist that were happily polymorphic
for the rearrangement (ie both rearranged and original versions of the chromosome
would exist in the gene pool). In fact, populations polymorphic for chromosomal
rearrangements are unknown in extant mammals, although separate populations
within the same species are commonly known to carry chromosomal rearrangements.

So, we have a real problem
with the simple model. On the one hand, if individuals heterozygous for
rearrangements are significantly partly sterile it is impossible for the rearrangement
to become established. On the other hand, if heterozygous individuals
are fertile, we should expect to see substantial rearrangement polymorphism
within populations, which we do not see, at least with mammals. And we
are stuck without a model for human/chimp lineage speciation as the allopatric
model seems inappropriate and the most likely sympatric model seems flawed.

The new model

This conundrum does have
a potential solution. A more recent model of population fitness in the
presence of chromosomal rearrangements points to the fact that recombination
is greatly reduced in individuals who are heterozygous for rearrangements. What
does this mean?

Well, recombination is a
process where, within meiosis (the cell division process that creates gametes,
eggs and
sperm, there is random mixing of genetic material in the pairs of chromosomes,
one derived from the father and one from the mother). Each individual
carries two copies of each chromosome in each of its somatic cells, one from
its mother and from its father. During meiosis, each pair of chromosomes
exchange material so that the chromosomes in the resulting sex cells (which
each have only one copy of each chromosome) contain a mixture of genetic material
from the father and the mother. This process of recombination or crossing over randomises the
genetic material and is a major contributor to genetic diversity.

Recombination is greatly
reduced in individuals that have one copy of a chromosome that is rearranged
and one that is not, specifically in those chromosomes that are rearranged.
Recombination of chromosomes heterozygous for a rearrangement is the main reason that heterozygous
individuals were held to be partly sterile, since recombination leads to substantial
deletions and duplications of genetic material resulting in unviable offspring.

But if there is little or
no recombination in chromosomes heterozygous for rearrangements, then heterozygous
individuals will be viable and fertile. There is no duplication or deletion
of genetic material, since there is no recombination.

Since individuals heterozygous
for rearrangements are fertile in the absence of recombination, rearrangements
will fix with the same probability as neutral mutations (assuming the rearrangement
is functionally neutral). And since there is no recombination in rearranged
chromosomes, there is a barrier to gene flow in those chromosomes. If
they do not recombine, then each karyotype (with and without the rearrangement)
can continue to exist in the population and can continue to interbreed with
viable offspring. However the mechanism for gene flow is blocked and each
version of the chromosome is entirely separate and can mutate with the same
degree of isolation as if the chromosome were isolated in separate species.

So we have a solution. Chromosomal
rearrangements occur as a result of random mutation. Recombination is
suppressed in those chromosomes allowing the mutation to fix in the population.
The suppression of recombination also isolates the genetic material on
that chromosome, and the chromosome evolves as though it were in a separate
non-interbreeding species. Over time more chromosomes mutate and become rearranged,
the lack of gene flow in the rearranged chromosomes leads to more and more divergent
evolution on those chromosomes eventually resulting in sexual incompatibility
between the populations that carry and do not carry the rearrangements and
speciation is complete.

Predictions

If it is true that chromosomal
rearrangements occurred before the speciation process was complete (or even
triggered it) and that
individuals with and without the rearrangement (and heterozygous hybrids) interbred
fertilely for some time, what signs would that leave in the genomes of human
and chimpanzee? Is there a smoking gun that we can seek as evidence
for this rather neat hypothesis.

There is. Remember that
during the period of interbreeding between the subpopulations, gene flow can
freely occur on chromosomes that have not undergone a rearrangement (these are
called co-linear chromosomes where the genes are found in the same order in
both subpopulations). Gene flow, however, is restricted between the subpopulations
on those chromosomes that have undergone a rearrangement (since there is no
recombination between the original and the rearranged karyotype). Therefore,
if there is a significant period when there is interbreeding between individuals
with and without the chromosomal rearrangement (and hybrids with both the original
and rearranged version of the chromosome), then one would expect to see a greater
degree of divergent evolutionary mutations on those chromosomes that had undergone
rearrangement compared with co-linear chromosomes. Why is this? Well,
since gene flow is not restricted in co-linear chromosomes, beneficial mutations
or neutral mutations linked to beneficial mutations through a selective sweep,
or other mutations that fix, will tend to fix in the entire population to the
same extent that they would do in any single species. On the other hand,
since there is no gene flow in the rearranged chromosomes, mutations that
fix in one version of the chromosome cannot spread to the other version and
so every fixed mutation in either case results in a difference (divergent
evolution) between original and rearranged chromosomes.

It is as though the co-linear
chromosomes are in a single fully interbreeding species, and the rearranged
chromosomes are already in separate non-breeding species.

Furthermore, if favourable
mutations which drive the divergence of the species accumulate on those chromosomes
that have rearrangements then that will lead to a signature on those chromosomes
indicating the action of positive selection. This is a key prediction.

Results

So, what do we see? In
a recent paper in Science (9), Navarro and Barton provide an answer. They
investigated rate of protein evolution on 115 autosomal genes across the chromosomes
of man and chimpanzee. The rate of protein evolution is given by a measure
for each gene, Ka/Ks, where Ka is the number
of non-synonymous single nucleotide substitutions between the human and
chimpanzee gene and Ks is the number of synonymous substitutions.
A synonymous substitution is one which codes for the same amino acid,
and hence protein and which has no effect on the amino acid, protein or organism
and is therefore not under selection. (Remember that there are 64 possible
combinations of 3 base pair codons, coding for only 20 amino acids. There is
therefore coding redundancy and each amino acid is coded for by more than one
codon - so some substitution mutations are silent and have no consequence in
the protein. These are synonymous mutations). A non-synonymous mutation
results in a different amino acid and protein and has a potential consequence
for the organism. Non-synonymous mutations, are therefore subject to selection.

First of all, the average
Ks across all the genes is 1.53% and the average Ka is 0.76% (ie,
in genes, only 1 in 150 nucleotides are non-synonymously different between man
and chimp: or in other words only 1 in 50 amino acids differ) - this is
well within the range of known coding divergence between chimpanzee and humans

Critically, the Ka/Ks ratio
for rearranged chromosomes was 0.84 and for co-linear chromosomes was 0.37.
This is highly significant and indicates that protein divergence is much
greater in the rearranged chromosomes than in the co-linear chromosomes exactly
as predicted. Furthermore if we look at the number of genes with
a Ka/Ks ratio > 1, a measure that indicates positive
selection at that gene, we find that genes on rearranged chromosomes have Ka/Ks >
1 in about 40% of cases whereas genes on co-linear chromosomes have Ka/Ks >
1 in only 8% of cases.

Navarro and Barton consider
many potential confounding factors for these data, but find none.

They also find evidence
of reduced flow of genes in rearranged chromosomes, as the hypothesis
would predict. In two sets of genes selected to measure absolute
K, the number of substitutions per 100 nucleotides, they find that K is greater
in rearranged than in co-linear chromosomes as predicted.

In addition, a reduced
rate of neutral polymorphisms in rearranged chromosomes is found. This
supports the hypothesis that speciation involved the accumulation of beneficial
mutations on the rearranged chromosomes, since the process of fixing those mutations
would have resulted in selective sweeps that reduced the rate of neutral polymorphism
on linked sites.

Conclusion

So the evidence is quite
strong that the hypothesis is correct. The concept that chromosomal rearrangements initially
led to chromosomal polymorphism within the breeding population. Gene flow
would not occur across the original and rearranged types of the chromosome resulting
in a greater evolutionary divergence on those chromosomes than co-linear
ones. Indeed, those chromosomes would become a focus for beneficial mutations
which were incompatible, driving the divergence of the populations and eventually
resulting in total reproductive isolation and speciation. The signature
of this is the greater accumulation of protein modifications on genes on rearranged
chromosomes. Indeed a high (40%) percentage of genes on rearranged chromosomes
are under positive selection as defined by Ka/Ks >1.

There are, of course, as
Navarro and Barton point out, other possible hypotheses to explain the data
including that rearrangement facilitates fixation or that rearrangements result
in relaxation of purifying selection and so on. However, the data does support
the hypothesis that chromosomal rearrangements fix by the period of post-rearrangement
hybridisation and that rearranged chromosomes are a focus for divergent mutations
which both trigger and ultimately complete the speciation.

We look forward to the publication
of the draft chimpanzee genome as a great deal of light will be shed on this
and many other matters concerning the evolution of man.